Fig. 5.

End resection of each DNA tail by drNurA-HerA. A The 5′ end resection activity of drNurA-HerA has been analyzed using substrate O3. B The 3′ end resection activity of drNurA-HerA has been analyzed using substrate O4. 400 nM 5′ FAM-labeled substrate DNA (O3 or O4) was incubated with drNurA (4 μM protomer) and drHerA (12 μM protomer) in the presence of 2 mM MgCl₂ and 8 mM MnCl₂. Reactions were carried out in the absence or in the presence of 1 mM ATP, incubated at 37°C for 15, 30, or 45 min. Reactions were stopped by stop buffer, followed by boiling at 100°C for 5 min and flash-cooling on ice for 10 min. Products were analyzed on 15% denaturing TBE-PAGE and gels were imaged at FAM fluorescent mode. The types of generated products before denaturing have been annotated at the right side of the gel to aid the interpretation of the bands on the gels. Markers were created by mixing different lengths of 5′ FAM-labeled DNA oligos together. The cutting patterns of each hairpin substrate were shown at the top of each gel. Red area indicates these bases were linked by phosphorothioate bonds, which can protect this area of strands from digesting by nuclease. Arrows indicate putative cleavage sites in the presence of ATP